CEN1

TgCEN1 is a Ca2+-sensor centrin implicated in dynamic centrosome architecture and Ca2+-modulated target recognition in Toxoplasma gondii (PMID:32759683, PMID:36009009). It binds two Ca2+ ions with high and medium affinity and undergoes Ca2+-dependent conformational changes that expose hydrophobic patches characteristic of a Ca2+ sensor (PMID:32759683). TgCEN1 self-assembles in a Ca2+-, temperature-, and concentration-dependent manner through electrostatic and hydrophobic interactions, following an N-to-C mode in which the first 21 N-terminal residues of one molecule engage the C-lobe of another; deletion of these residues abolishes assembly, and the P17-XPC target peptide reduces it (PMID:32759683, PMID:34114596). Target recognition is lobe-specific: a high-affinity C-lobe site can constitutively engage target peptide while a low-affinity N-lobe site becomes active only upon Ca2+ stimulus (PMID:34114596). TgCEN1 directly binds distinct repeat motifs of the centrosomal protein TgSFI1 with differing affinities, Ca2+ sensitivities, and lobe-specific binding modes, so that its conformational plasticity underlies dynamic, Ca2+-regulated association with TgSFI1 at the centrosome (PMID:36009009). Cellular and in vivo functional roles beyond these in vitro biophysical characterizations have not been established in the available corpus.

1. 2020 Medium

   Whether TgCEN1 functions as a Ca2+ sensor was unresolved; demonstrating two-site Ca2+ binding with conformational exposure of hydrophobic patches established its sensor behavior and distinguished it functionally from TgCEN2.

   Evidence Ca2+ binding assays, CD, fluorescence spectroscopy, and dynamic light scattering on recombinant TgCEN1

   PMID:32759683

   Open questions at the time

   • No cellular localization or in vivo functional consequence tested
   • Physiological targets not yet identified
2. 2021 Medium

   How TgCEN1 self-associates was unknown; mapping self-assembly to an N-to-C interaction dependent on the first 21 residues defined the structural determinants of oligomerization and its modulation by Ca2+, temperature, and target peptide.

   Evidence DLS, ITC, NMR, CD, fluorescence spectroscopy, and N-terminal deletion mutant analysis

   PMID:34114596

   Open questions at the time

   • Functional role of self-assembly in vivo untested
   • Higher-order assembly state and stoichiometry at the centrosome unknown
3. 2021 Medium

   The mode of target-peptide recognition was undefined; dissecting binding into a constitutive high-affinity C-lobe site and a Ca2+-activated low-affinity N-lobe site established a two-site, differentially Ca2+-regulated recognition mechanism.

   Evidence ITC, NMR, CD, and fluorescence spectroscopy on intact TgCEN1 and isolated lobes titrated with P17-XPC peptide

   PMID:34114596

   Open questions at the time

   • P17-XPC is a model peptide, not a validated physiological target
   • Cellular relevance of lobe-specific binding not demonstrated
4. 2022 Medium

   A physiological binding partner was lacking; showing direct, lobe-specific, Ca2+-sensitive binding to distinct repeat motifs of TgSFI1 connected TgCEN1's biophysical properties to a defined centrosomal target.

   Evidence ITC, NMR, CD, and fluorescence spectroscopy using TgCEN1, isolated lobes, and synthetic TgSFI1 repeat peptides

   PMID:36009009

   Open questions at the time

   • Interaction shown with peptides in vitro, not full-length TgSFI1 in cells
   • Functional consequence for centrosome dynamics or parasite division untested

• The in vivo cellular function of TgCEN1 — its centrosomal localization, contribution to centrosome duplication or parasite division, and phenotype upon depletion — remains uncharacterized in the available corpus.
• No genetic knockout or knockdown phenotype
• No in-cell localization or division-cycle analysis
• Interaction with full-length TgSFI1 in the parasite not confirmed